Effect of maternal antibodies on induction and persistence of vaccine-induced immune responses against bovine viral diarrhea virus type II in young calves.

OBJECTIVE: To determine the effect of maternally derived antibodies on induction of protective immune responses against bovine viral diarrhea virus (BVDV) type II in young calves vaccinated with a modified-live bovine viral diarrhea virus (BVDV) type I vaccine. DESIGN: Blinded controlled challenge study. ANIMALS: 24 neonatal Holstein and Holstein-cross calves that were deprived of maternal colostrum and fed pooled colostrum that contained a high concentration of (n = 6) or no (18) antibodies to BVDV. PROCEDURE: At 10 to 14 days of age, 6 seropositive and 6 seronegative calves were given a combination vaccine containing modified-live BVDV type I. All calves were kept in isolation for 4.5 months. Six calves of the remaining 12 untreated calves were vaccinated with the same combination vaccine at approximately 4 months of age. Three weeks later, all calves were challenged intranasally with a virulent BVDV type II. RESULTS: Seronegative unvaccinated calves and seropositive calves that were vaccinated at 2 weeks of age developed severe disease, and 4 calves in each of these groups required euthanasia. Seronegative calves that were vaccinated at 2 weeks or 4 months of age developed only mild or no clinical signs of disease. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicate that a single dose of a modified-live BVDV type-I vaccine given at 10 to 14 days of age can protect susceptible young calves from virulent BVDV type II infection for at least 4 months, but high concentrations of BVDV-specific maternally derived antibodies can block the induction of the response.

Association of porcine circovirus 2 with reproductive failure in pigs: a retrospective study, 1995-1998.

In order to determine if vertically transmitted porcine circovirus (PCV) has played a role in reproductive failure in pigs in areas of endemic infection, archival fixed tissues were examined by polymerase chain reaction (PCR) and immunohistochemistry. Tissues tested were from routine cases of abortion or reproductive failure submitted between 1995 and 1998 to the diagnostic laboratory at the Western College of Veterinary Medicine, Saskatoon. They originated from 29 high-health herds in the provinces of Alberta and Saskatchewan and comprised a total of 36 individual submissions. Porcine circovirus type 1 (PCV1) was not detected by PCR in any submitted tissues. Porcine circovirus type 2 (PCV2) was not detected by PCR or immunohistochemistry in any of the submitted tissue. The effect of extended formalin fixation on the detection of PCV2 by PCR was assessed and fixation for up to one week had no gross effect on sensitivity of detection using this PCR technique. Failure to detect porcine circoviruses in cases of reproductive failure prior to 1999 in areas of endemic infections, suggests that reproductive disease may be a new clinical manifestation of PCV2 infection, and that vertical transmission may not have been the primary mechanism of initial dissemination of the virus in the pig population.

Multiple porcine circovirus 2-associated abortions and reproductive failure in a multisite swine production unit.

Porcine circovirus type 2 was detected in several stillborn and nonviable neonatal piglets presenting with chronic passive congestion, cardiac hypertrophy, and severe diffuse myocarditis. The presence of the virus in the heart and other tissues of affected piglets was confirmed by polymerase chain reaction, immunohistochemistry, and virus isolation techniques. Other reproductive losses and associated infectious agents in the herd are discussed.

Lack of antibodies to porcine circovirus type 2 virus in beef and dairy cattle and horses in western Canada.

Porcine circovirus type 2 (PCV2) is a recently recognized agent that is consistently associated with postweaning multisystemic wasting disease in swine. There are conflicting data concerning the ability of this virus to infect and cause disease in other species. To determine if normal cattle, cattle affected with various illnesses, and normal horses in endemic areas of PCV2 infection in swine have had PCV2 infections, 100 randomly selected bovine sera, 100 equine sera, and 100 colostrum samples from clinically normal dairy cattle were examined for the presence of antibodies to porcine circoviruses by using ELISAs. All samples tested were negative for antibodies to porcine circoviruses. As well, a seronegative neonatal Holstein calf and 6 seronegative, 6-month-old beef calves that were experimentally infected with PCV2 failed to develop antibodies to the virus. These results suggest that natural infection of cattle and horses with PCV2 does not occur, or is a rare event, in western Canada.

Efficacy of an inactivated respiratory syncytial virus vaccine in calves.

OBJECTIVE: To determine whether an inactivated bovine respiratory syncytial virus (BRSV) vaccine would protect calves from infection with virulent BRSV. DESIGN: Randomized controlled trial. ANIMALS: 27 nine-week-old calves seronegative for BRSV exposure. PROCEDURE: Group-1 calves (n = 9) were not vaccinated. Group-2 calves (n = 9) were vaccinated on days 0 and 21 with an inactivated BRSV vaccine containing a minimum immunizing dose of antigen. Group-3 calves (n = 9) were vaccinated on days 0 and 21 with an inactivated BRSV vaccine containing an amount of antigen similar to that in a commercial vaccine. All calves were challenged with virulent BRSV on day 42. Clinical signs and immune responses were monitored for 8 days after challenge. Calves were euthanatized on day 50, and lungs were examined for lesions. RESULTS: Vaccination elicited increases in BRSV-specific IgG and virus neutralizing antibody titers and in production of interferon-gamma. Virus neutralizing antibody titers were consistently less than IgG titers. Challenge with BRSV resulted in severe respiratory tract disease and extensive pulmonary lesions in control calves, whereas vaccinated calves had less severe signs of clinical disease and less extensive pulmonary lesions. The percentage of vaccinated calves that shed virus in nasal secretions was significantly lower than the percentage of control calves that did, and peak viral titer was lower for vaccinated than for control calves. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that the inactivated BRSV vaccine provided clinical protection from experimental infection with virulent virus and decreased the severity of pulmonary lesions. Efficacy was similar to that reported for modified-live BRSV vaccines.

Effect of vaccination on experimental infection with Bordetella bronchiseptica in dogs.

OBJECTIVE: To determine comparative efficacy of vaccines administered IM and intranasally, used alone or sequentially, to protect puppies from infection with Bordetella bronchiseptica and determine whether systemic or mucosal antibody response correlated with protection. DESIGN: Randomized controlled trial. ANIMALS: 50 specific-pathogen-free Beagle puppies. PROCEDURE: In 2 replicates of 25 dogs each, 14-week-old puppies that were vaccinated against canine distemper virus and parvovirus were vaccinated against B bronchiseptica via intranasal, IM, intranasal-IM, or IM-intranasal administration or were unvaccinated controls. Puppies were challenge exposed via aerosol administration of B bronchiseptica 2 weeks after final vaccination. Clinical variables and systemic and mucosal antibody responses were monitored for 10 days after challenge exposure. Puppies in replicate 1 were necropsied for histologic and immunohistochemical studies. RESULTS: Control puppies that were seronegative before challenge exposure developed paroxysmal coughing, signs of depression, anorexia, and fever. Vaccinated puppies (either vaccine) that were seronegative before challenge exposure had fewer clinical signs. Puppies that received both vaccines had the least severe clinical signs and fewest lesions in the respiratory tract. Vaccinated dogs had significantly higher concentrations of B bronchiseptica-reactive antibodies in serum saliva before and after challenge. Antibody concentrations were negatively correlated with bacterial growth in nasal cavity and pharyngeal samples after challenge exposure. CONCLUSIONS AND CLINICAL RELEVANCE: Parenterally and intranasally administered vaccines containing B bronchiseptica may provide substantial protection from clinical signs of respiratory tract disease associated with infection by this bacterium. Administration of both types of vaccines in sequence afforded the greatest degree of protection against disease.

Development and application of a competitive enzyme-linked immunosorbent assay for the detection of serum antibodies to porcine circovirus type 2.

We report the development of a competitive enzyme-linked immunosorbent assay (c-ELISA) for the detection of antibodies to porcine circovirus type 2 (PCV2), the agent associated with the recently described postweaning multisystemic wasting syndrome in pigs. At present, no method has been published describing a c-ELISA for the detection of antibodies to PCV2, and currently employed tests are impractical for use in some laboratories. The assay described here uses a cell culture isolate of porcine circovirus type 2 as antigen and a PCV2-specific monoclonal antibody as the competing reagent. Evaluation of the ELISA was performed by comparison with results obtained using an indirect immunofluorescent test on 484 sera from pig herds in the United Kingdom, Canada, France, and the USA and serial bleeds from pigs experimentally infected with porcine circoviruses. The sensitivity and specificity of the ELISA were determined as 99.58% and 97.14%, respectively, at 2 standard deviations (SD) from the mean or 95.81% and 100% at 3 SD from the mean. Using this ELISA, a serologic survey of 461 sera collected from commercial pig herds in Northern Ireland between 1973 and 1999 was undertaken. Analysis of the results of this survey demonstrated that the number of ELISA-positive sera detected in an individual year during this period ranged from 55% to 100%. This c-ELISA has applications for large-scale rapid diagnosis of PCV2 infection in pig populations worldwide and for immunoscreening of sera from other species for antibodies to PCV2.

Absence of evidence for porcine circovirus type 2 in cattle and humans, and lack of seroconversion or lesions in experimentally infected sheep.

No antibodies to porcine circovirus type 2 were detected in sera from cattle, sheep and humans. Experimental infection of lambs with this virus failed to produce lesions or seroconversion.

Coinfection by porcine circoviruses and porcine parvovirus in pigs with naturally acquired postweaning multisystemic wasting syndrome.

Postweaning multisystemic wasting syndrome (PMWS) is an emerging disease in swine. Recently, the disease has been reproduced with inocula containing a newly described porcine circovirus (PCV), designated PCV 2, and porcine parvovirus (PPV). In order to determine if these viruses interact in naturally acquired PMWS, affected tissues from field cases were examined by immunohistochemistry (IHC) and polymerase chain reaction (PCR) for PCV 2 and PPV, as well as by PCR for the other recognized porcine circovirus, PCV 1. Porcine circovirus 2 was detected by PCR or IHC in affected fixed or frozen tissues from 69 of 69 cases of PMWS collected over 3 years from 25 farms. Porcine parvovirus was detected in 12 of the same cases, and PCV 1 was detected in 9 of 69; however, an apparent decrease was found in the sensitivity of the PCRs used to detect the latter 2 viruses when fixed tissue from the same cases were compared with the use of frozen tissues. Porcine circovirus 2 was not detected by PCR in affected tissues from 16 age-matched pigs that had Streptococcus suis-associated disease. Electron microscopic examination of plasma pooled from 15 pigs with PMWS revealed the presence of PCV and PPV, whereas these viruses were not observed in pooled plasma from 5 age-matched clinically normal pigs. These results confirm and extend previous findings documenting a consistent association of PCV 2 with PMWS. As well, infection by PPV or PCV 1 or both may be an important cofactor in the pathogenesis of some, but apparently not all, cases of PMWS.

The efficacy of modified-live bovine respiratory syncytial virus vaccines in experimentally infected calves.

The efficacy of modified-live (MLV) bovine respiratory syncytial virus (BRSV) vaccines and the correlates of vaccine-induced immunity were investigated in calves using a virulent experimental infection. Clinical disease and pulmonary pathology were significantly reduced, relative to unvaccinated controls, in calves vaccinated according to label directions with commercial multivalent MLV BRSV vaccines. In vitro assays of cellular immunity were more consistent correlates of vaccine associated protection than presence of post vaccination serum antibody. Most vaccinated calves shed virus, but peak virus titre was suppressed compared to unvaccinated controls, with clearance coincident with the simultaneous appearance of mucosal antibody, cytotoxic cells in the lung and anamnestic or primary serum antibody responses. Virus clearance in unvaccinated calves was coincident with the appearance of BRSV specific cytotoxic cells, before mucosal antibody was detected.

Lack of evidence of conserved lentiviral sequences in pigs with post weaning multisystemic wasting syndrome.

In order to investigate the role of retroviruses in the recently described porcine postweaning multisystemic wasting syndrome (PMWS) serum and leukocytes were screened for reverse transcriptase (RT) activity, and tissues were examined for the presence of conserved lentiviral sequences using degenerate primers in a polymerase chain reaction (PCR). Serum and stimulated leukocytes from the blood and lymph nodes from pigs with PMWS, as well as from control pigs had RT activity that was detected by the sensitive Amp-RT assay. A 257-bp fragment was amplified from DNA from the blood and bone marrow of pigs with PMWS. This fragment was identical in size to conserved lentiviral sequences that were amplified from plasmids containing DNA from several lentiviruses. Cloning and sequencing of the fragment from affected pigs, however, did not reveal homology with the recognized lentiviruses. Together the results of these analyses suggest that the RT activity present in tissues from control and affected pigs is the result of endogenous retrovirus expression, and that a lentivirus is not a primary pathogen in PMWS.

The effect of formalin-inactivated vaccine on respiratory disease associated with bovine respiratory syncytial virus infection in calves.

The effect of vaccination with a formalin-inactivated, alum-precipitated (FI), bovine respiratory syncytial virus (BRSV) vaccine on BRSV induced respiratory disease in calves was investigated. Six month old BRSV-naive calves were vaccinated with either a FI, a modified live virus (MLV), or virus antigen negative control vaccine (n = 4 per group). One month after the second vaccination, the calves were aerosol challenged with lung wash from a newborn calf infected with a field isolate of BRSV. Moderate to severe clinical disease occurred in all calves. Calves that received FI vaccine had a significantly earlier (day 2 vs. day 4-5) onset of pyrexia and dyspnea (P < 0.05). Pulmonary lesions, consisting of cranioventral atelectasis and dorsal emphysema, occurred in all groups. Two calves that received MLV, and three that received FI vaccine, had reduced pneumonic lung area relative to controls. Vaccination with the FI vaccine resulted in more rapid onset of clinical disease, but ultimately, reduced pulmonary pathology in most recipients.

Reproduction of lesions of postweaning multisystemic wasting syndrome in gnotobiotic piglets.

Neonatal gnotobiotic piglets were inoculated with tissue homogenates and low- and high-passage cell culture material to determine if the lesions of the newly described porcine postweaning multisystemic wasting syndrome (PMWS) could be reproduced. For this, 17 3-day-old gnotobiotic piglets were inoculated intranasally with pelleted chloroform-treated, filtered extracts from cell cultures, filter-sterilized homogenates of lymphoid tissue from PMWS-affected piglets, or control materials. Piglets were maintained in germ-free isolators for up to 5 weeks after infection prior to euthanasia and collection of samples for analysis. All piglets inoculated with the viral inocula developed lesions typical of PMWS, including generalized lymphadenopathy, hepatitis, nephritis, interstitial pneumonia, myocarditis, and gastritis. Porcine circovirus (PCV), as well as porcine parvovirus (PPV), was detected in tissues by virus reisolation, polymerase chain reaction analysis, or immunohistochemistry. All infected piglets developed moderate to high titers of antibody to PCV and moderate titers to PPV. No lesions, virus, or virus-specific antibodies were detected in sham-inoculated or uninoculated control piglets. These studies demonstrate that the lesions of PMWS can be experimentally reproduced in gnotobiotic piglets using filterable viral agents derived from pigs with PMWS and provide an experimental basis for further investigation into the pathogenesis and control of this emerging infectious disease in swine.

Myocarditis and abortion associated with intrauterine infection of sows with porcine circovirus 2.

Porcine circovirus 2 (PCV2) is a recently identified agent that has been associated with postweaning multisystemic wasting syndrome (PMWS) in swine populations. In this report, the potential spectrum of disease associated with PCV2 is expanded by evidence of vertical transmission and associated reproductive failure. PCV2 was isolated from a litter of aborted piglets from a farm experiencing late-term abortions and stillbirths. Severe, diffuse myocarditis was present in 1 piglet associated with extensive immunohistochemical staining for PCV2 antigen. Variable amounts of PCV2 antigen were also present in liver, lung, and kidney of multiple fetuses. The presence of other agents that have been associated with fetal lesions and abortion in swine, including porcine parvovirus, porcine reproductive respiratory syndrome virus, encephalomyocarditis virus, and enterovirus, could not be established.